Ischemic brain damage leads to cognitive deficits, long-term disability, and even death. This damage occurs when the blood supply to the brain is arrested, which takes place during cardiac surgeries, stroke, or myocardial infarction. NMDA channel blockers and other drugs designed to protect the brain from ischemic damage have failed in clinical trials. Therefore, the need to identify novel targets for pharmacological intervention during brain ischemia is very urgent. Our preliminary data indicate that we may be able to identify such novel target(s). The toxic phase of brain ischemia is executed at the time when neurons are profoundly depolarized by highly elevated extracellular K+ ([K+]o). This [K+]o elevation is caused by K+ efflux from the cytosol to the extracellular medium and coincides with a reciprocal Na+ influx. As a result, ischemic neurons are depolarized and contain elevated cytosolic Na+. In preliminary work, we simulated such ischemic depolarization in cultured neurons and found that cytosolic [Ca2+] ([Ca2+]c) elevations resulted from an Na-dependent Ca2+ influx that could not be prevented by blocking NMDA channels. Interestingly, the Na-dependent [Ca2+]c elevations activated a pathway of potentially even more neurotoxic Zn2+ influx. The Na-dependent [Ca2+]c elevations in depolarized ischemic neurons are likely mediated by plasmalemmal K-dependent Na+/Ca2+ exchangers, NCKXs. Since NCKX2 is a major NCKX isoform expressed in the brain cortex, we would like to explore the idea that there is a causal link between NCKX2-mediated Ca2+ influx and an activation of Zn2+ influx. To test this idea, we plan to use cultured cortical neurons obtained from wild type and NCKX2 knockout mice. Our project has 2 specific aims and will be completed in 2 years. In Aim 1, we will determine the impact of the NCKX2 knockout on the rate of cytosolic Zn2+ ([Zn2+]c) elevation in ischemic neurons. In Aim 2, we will determine the impact of the NCKX2 knockout and Zn2+ on ischemic neuronal death. If we find that NCKX2 operation is causally linked to the activation of neurotoxic Zn2+ influx in ischemic neurons, our data will provide support for developing inhibitors of Zn2+influx activation by NCKX2. Such agents may become neuroprotective drugs that could be used to improve the safety of cardiac surgeries. PUBLIC HEALTH RELEVANCE: About 32% of the very commonly performed cardiac surgeries involving cardiopulmonary bypass (CPB) cause ischemic events in the brain and carry a significant risk of post-operative cognitive deficits. This project explores the idea that the brain damage relevant to CPB is caused by the abnormal operation of a K- dependent Na+/Ca2+ exchanger that activates a neurotoxic Zn2+ influx. This research may lead to the development of a novel therapy that could be applied prior to CPB surgeries to improve their safety.